Rapid genetic screening with high quality factor metasurfaces

Genetic analysis methods are foundational to advancing personalized and preventative medicine, accelerating disease diagnostics, and monitoring the health of organisms and ecosystems. Current nucleic acid technologies such as polymerase chain reaction (PCR), next-generation sequencing (NGS), and DNA microarrays rely on fluorescence and absorbance, necessitating sample amplification or replication and leading to increased processing time and cost. Here, we introduce a label-free genetic screening platform based on high quality (high-Q) factor silicon nanoantennas functionalized with monolayers of nucleic acid fragments. Each nanoantenna exhibits substantial electromagnetic field enhancements with sufficiently localized fields to ensure isolation from neighboring resonators, enabling dense biosensor integration. We quantitatively detect complementary target sequences using DNA hybridization simultaneously for arrays of sensing elements patterned at densities of 160,000 pixels per cm2. In physiological buffer, our nanoantennas exhibit average resonant quality factors of 2,200, allowing detection of two gene fragments, SARS-CoV-2 envelope (E) and open reading frame 1b (ORF1b), down to femtomolar concentrations. We also demonstrate high specificity sensing in clinical nasopharyngeal eluates within 5 minutes of sample introduction. Combined with advances in biomarker isolation from complex samples (e.g., mucus, blood, wastewater), our work provides a foundation for rapid, compact, amplification-free and high throughput multiplexed genetic screening assays spanning medical diagnostics to environmental monitoring.


Supplementary Note 2: Spatial distribution of electric fields around resonators
The sensitivity of a resonant mode to minute changes in the local refractive index can be estimated by the fraction of electric field energy residing outside the resonator. We calculate the exposure of the mode utilized in our sensors with the following equation: where out and in are the permittivity of the medium containing the analyte and the permittivity of the resonator and substrate, respectively. V out and V in represent the volumetric regions of the analyte containing medium and the portions inside the resonator or substrate that do not overlap with any bound materials or molecules. Performing this analysis on the sensor design described in the main text as well as guided mode resonant structures previously described in reference [1] composed of notched silicon waveguides, we find that our silicon block chains significantly increase field penetration into the surrounding environment. Field profiles of the two structures are plotted in Supplementary Fig. 4 showing similar transverse electric waveguide modes. Due to the subwavelength spacing of the discrete silicon blocks in our sensors, we still excite the localized waveguide modes along the periodic direction that are seen in continuous silicon wire waveguides. However, the grating-like structure exposes regions of the mode to the surroundings while also reducing the effective mode index of the waveguide, leading to further extension of the fields out of the resonator. This design results in the fraction of the mode energy in the surroundings to increase to f U E = 0.29 compared to only f U E = 0.08 for notched or continuous waveguide structures.

Supplementary Note 3: Quality factor scaling and water absorption
As discussed in the main text, introducing an asymmetry along a silicon waveguide allows for the excitation of previously bound modes. Reduction of the asymmetry, ∆w in the case of our metasurfaces, decreases the coupling strength of the mode to free-space radiation thereby increasing the Q factor. For a material that exhibits no intrinsic absorption losses, such as silicon in the near infrared, the Q factor can be arbitrarily increased as the perturbation strength approaches zero. This dependence of the Q factor on subtle structural deviations have been previously described through temporal coupled-mode theory and perturbation theory [2,3,4]: where B is a constant that depends on the resonator geometry and α is a unit-less asymmetry parameter represented by ∆w/w 0 in our metasurface. This relationship is shown in Supplementary  Fig. 5, where theory (solid line) and numerical simulations (stars) indicate diverging Q factors as ∆w is decreased. We also observe that experimentally observed Q factors are lower than predicted values (experimental data from Main text Fig. 2). One significant factor limiting our experimental quality factors is the absorption coefficient of water at telecommunication wavelengths. Since all our optical measurements are performed in aqueous solutions, dissipative losses are expected to decrease our measured Q factors as shown by the dashed line in Supplementary Fig. 5, which represents numerical calculations including water absorption. The effects of absorption losses are particularly strong as ∆w is decreased, as longer resonance lifetimes lead to greater interaction between the resonant mode and the absorptive background medium. Future iterations of our sensor can be designed in the water absorption window around 1300 nm to maximize performance of the resonators. Additionally, fabrication imperfections such as surface roughness or non-uniformity in the metasurface structures will introduce scattering losses and reduce the observed Q factor.

Supplementary Note 4: Finite size resonators
While the resonators shown in the main text exhibit high-Q modes in longer 1-D arrays (200 µm), we show that the resonators can be scaled down significantly while maintaining sharp spectral features. Our metasurface design features low scattering losses out the ends of the waveguides, and hence are relatively robust to resonator finite size effects due to the high index contrast between separated silicon blocks and gaps containing the background medium. In Supplementary Fig. 6a, we show calculated dispersion diagrams for three different resonators consisting of a solid silicon waveguide with increasing depths of notch corrugations. The waveguide has width of 600 nm and from top to bottom, the bands correspond to notches added on both sides of the waveguide with depths of 50, 150, and 300 nm. We observe flattening of the bands as the notch depth is increased until 300 nm, where the waveguide is now separated into distinct silicon blocks. The flatter bands indicate a much smaller group velocity due to strong in-plane Bragg scattering, which reduces the propagation of the mode out the waveguide ends and reduces effects of shrinking the resonator on the Q factor. We experimentally verify that we can maintain high quality factors while shortening the overall length of each resonator. In Supplementary Fig. 6b,c we show SEM images of multiple resonators with varying lengths from 300 µm down to 50 µm and representative spectra. Fitting N=6 resonators for each condition, Supplementary Fig. 6d shows little change in the Q factor with varying waveguide length. Resonators with ∆w = 50 and 30 nm maintain high Q factors exceeding 1000 even in resonators down to 50 µm. Each resonator could potentially be further scaled down with added dielectric mirrors patterned on the waveguide ends to reduce scattering losses. Thus, it is possible to envision individual free space coupled high Q resonators on the order of a few µm.
where ∆λ/∆n s the resonant wavelength shift induced by a change in the background medium refractive index or bulk refractive index sensitivity, Q is the resonator quality factor, and λ 0 is the resonant wavelength. To determine the bulk refractive index sensitivity of our devices, we take a series of optical measurements in various saline solutions with differing concentrations of NaCl dissolved in deionized water. Increasing NaCl concentrations have been shown to increase the refractive index of water. [5] In Supplementary Fig.7, the change in resonant wavelength indicates our sensors have a sensitivity of ∆λ/∆n = 270 nm/RIU. The sensitivity of the resonators does not vary significantly with ∆w, as the block asymmetry alters the Q factor, but the modal overlap with the surrounding medium does not change. Given our resonator Q factors of ∼2200 (Main text Fig.  2), we obtain a sensing FOM of around 400. This value is larger than previous demonstrations in plasmonic or dielectric metasurface based sensors. [6,7,8,9,10] We also note that modification of our metasurfaces, such as introducing subwavelength gaps or slots along the waveguide that further expose the resonant mode to target analytes, could dramatically improve the sensitivity and FOM of future iterations of the devices.

Supplementary Note 6: Modeling self-assembled monolayers
To estimate the resonant wavelength shifts corresponding to successive molecular layers deposited on our sensors (Main text Fig. 3c), we model each surface step with a thin dielectric shell and numerically calculate the response with FDTD as described above. The dielectric shells extend from all exposed silicon faces of our metasurface nanoblocks as shown in Supplementary Fig. 8. The initial bare sensor before surface functionalization is calculated with a 4 nm silicon dioxide layer due to the thermal passivation step performed after nanofabrication of our sensors. [11] The AUTES layer is assumed to bind as a uniform monolayer of thickness 1.8 nm and refractive indices ranging from n = 1.40-1.45, based on reported literature values. The refractive index is calculated for a range that corresponds to typical estimated optical properties of biomolecular layers. The MBS layer is estimated as a 0.7 nm thick layer based on the reported spacer arm length of the molecule and refractive index of n = 1.40-1.45. The thiolated ssDNA probe layers are calculated with optical properties n = 1.37-1.382, accounting for potential reduction in density due to dilution with PEG chains. [12,13,14] We estimate the structure of our single-stranded DNA probes using the open source software "mfold" to predict the secondary structure in a 1X PBS solution and the most stable conformation is shown in Supplementary Fig. 9. The probe layer thickness is estimated by the wormlike chain model [15]: where the monomer spacing a = 0.6 nm, persistence length L p = 1 nm, and contour length l = Na (N = number of nucleotides) give a radius of gyration, Rg, and layer thickness of ∼ 4 nm. [16] The dsDNA probe refractive index is approximated as n = 1.4-1.423 as estimated from the densification and increased polarizability of duplexed DNA compared to single stranded DNA. [17] The thickness is similarly estimated by the wormlike chain model, but with a = 0.3 nm and Lp = 30 nm, which returns a similar radius of gyration of 4 nm. Based on the short fragment length of our probe and targets at 22-26 nt, we do not expect a significant monolayer thickness change upon DNA hybridization.

Supplementary Note 7: Fluorescence microscopy
Fluorescence experiments were performed after DNA hybridization experiments with target nucleic acids tagged with ATTO590 dye on the 5' end. Dried samples were placed in a Zeiss Ax-ioImager system and imaged with a 20x objective. Fluorescence images were acquired with 1000 ms exposures on a Zeiss Axiocam 506 mono camera. Fluorescence intensity values were averaged over a 80 x 40 µm area and were normalized to the maximum intensity values from chips hybridized with complementary E gene targets as seen in Supplementary Fig. 10.

Supplementary Note 8: Langmuir adsorption model
Concentration dependent resonant wavelength shift responses in main text Fig. 4 were fit to the Hill equation (formally equivalent to the Langmuir isotherm): where θ max is the saturated maximum binding signal at high target concentrations, X is the target concentration, h is the Hill coefficient which describes the slope of the curve, and K d is the concentration value that corresponds to half-maximum binding signals. Furthermore, the Langmuir adsorption model is also used to fit the time varying responses in main text Fig. 5. The wavelength shift response as a function of time is described as [18]: where θ eq is the saturated equilibrium binding signal and k is an "observed" rate constant that accounts for both target hybridization and reversible dehybridization rates.
Supplementary Fig. 1. Schematic of near-infrared microscope set up utilized to collect spectra from metasurface samples.
Supplementary Fig. 2. Multi-step surface functionalization of a DNA probe monolayer on silicon metasurfaces.
Supplementary Fig. 3. a, Simulated waveguide dispersion in an unperturbed chain of subwavelength silicon blocks. b, Brillouin zone folding introduced via symmetry breaking in biperiodic guided mode resonator.
Supplementary Fig. 4. a, Electric field profile for notched silicon waveguide on a sapphire substrate. Upper panel shows an x-y cut through the center of the structure and the lower panel is an x-z cut through the center of the notch perturbation where fields are most strongly concentrated. b, Electric field profile for asymmetric chain of silicon blocks on a sapphire substrate. Upper panel represents the x-y cut through the center of the structure and the lower panel is an x-z cut through the center of the smaller block. Scale bar is 200 nm. Supplementary Fig. 5. Resonator quality factor as a function of difference in neighboring silicon block length, ∆d.
Supplementary Fig. 6. a, Simulated waveguide dispersion in waveguides with varying notch depth. b,Representative optical spectra from waveguides with length of 300 µm and 50 µm. c, SEM image of resonators fabricated with different waveguide lengths. d, Quality factors as a function of waveguide length. Supplementary Fig. 9. Likely secondary structure of DNA probes used in this work. Probes sequences were optimized to avoid formation of stable loop structures.  Table 1. Probe and target sequences used in this work.